From hazard to risk prioritization: a case study to predict drug-induced cholestasis using physiologically based kinetic modeling.

Cholestasis is characterized by hepatic accumulation of bile acids. Clinical manifestation of cholestasis only occurs in a small proportion of exposed individuals. The present study aims to develop a new approach methodology (NAM) to predict drug-induced cholestasis as a result of drug-induced hepatic bile acid efflux inhibition and the resulting bile acid accumulation. To this end, hepatic concentrations of a panel of drugs were predicted by a generic physiologically based kinetic (PBK) drug model. Their effects on hepatic bile acid efflux were incorporated in a PBK model for bile acids. The predicted bile acid accumulation was used as a measure for a drug's cholestatic potency. The selected drugs were known to inhibit hepatic bile acid efflux in an assay with primary suspension-cultured hepatocytes and classified as common, rare, or no for cholestasis incidence. Common cholestasis drugs included were atorvastatin, chlorpromazine, cyclosporine, glimepiride, ketoconazole, and ritonavir. The cholestasis incidence of the drugs appeared not to be adequately predicted by their Ki for inhibition of hepatic bile acid efflux, but rather by the AUC of the PBK model predicted internal hepatic drug concentration at therapeutic dose level above this Ki. People with slower drug clearance, a larger bile acid pool, reduced bile salt export pump (BSEP) abundance, or given higher than therapeutic dose levels were predicted to be at higher risk to develop drug-induced cholestasis. The results provide a proof-of-principle of using a PBK-based NAM for cholestasis risk prioritization as a result of transporter inhibition and identification of individual risk factors.

Population pharmacokinetic model to generate mechanistic insights in bile acid homeostasis and drug-induced cholestasis.

Bile acids (BA) fulfill a wide range of physiological functions, but are also involved in pathologies, such as cholestasis. Cholestasis is characterized by an intrahepatic accumulation of BAs and subsequent spillage to the systemic circulation. The aim of the present study was to develop physiologically based kinetic (PBK) models that would provide a tool to predict dose-dependent BA accumulation in humans upon treatment with a Bile Salt Export Pump (BSEP) inhibitor. We developed a PBK model describing the BA homeostasis using glycochenodeoxycholic acid as an exemplary BA. Population wide distributions of BSEP abundances were incorporated in the PBK model using Markov Chain Monte Carlo simulations, and alternatively the total amount of BAs was scaled empirically to describe interindividual differences in plasma BA levels. Next, the effects of the BSEP inhibitor bosentan on the BA levels were simulated. The PBK model developed adequately predicted the in vivo BA dynamics. Both the Markov Chain Monte Carlo simulations based on a distribution of BSEP abundances and empirical scaling of the total BA pool readily described the variations within and between data in human volunteers. Bosentan treatment disproportionally increased the maximum BA concentration in individuals with a large total BA pool or low BSEP abundance. Especially individuals having a large total BA pool size and a low BSEP abundance were predicted to be at risk for rapid saturation of BSEP and subsequent intrahepatic BA accumulation. This model provides a first estimate of personalized safe therapeutic external dose levels of compounds with BSEP-inhibitory properties.

Intestinal in vitro transport assay combined with physiologically based kinetic modeling as a tool to predict bile acid levels in vivo.

Bile acid homeostasis is vital for numerous metabolic and immune functions in humans. The enterohepatic circulation of bile acids is extremely efficient, with ~95% of intestinal bile acids being reabsorbed. Disturbing intestinal bile acid uptake is expected to substantially affect intestinal and systemic bile acid levels. Here, we aimed to predict the effects of apical sodium-dependent bile acid transporter (ASBT)-inhibition on systemic plasma levels. For this, we combined in vitro Caco-2 cell transport assays with physiologically based (PBK) modeling. We used the selective ASBT-inhibitor odevixibat (ODE) as a model compound. Caco-2 cells grown on culture inserts were used to obtain transport kinetic parameters of glycocholic acid (GCA). The apparent Michaelis-Menten constant (Km,app), apparent maximal intestinal transport rate (Vmax,app), and ODE's inhibitory constant (Ki) were determined for GCA. These kinetic parameters were incorporated into a PBK model and used to predict the ASBT inhibition effects on plasma bile acid levels. GCA is transported over Caco-2 cells in an active and sodium-dependent manner, indicating the presence of functional ASBT. ODE inhibited GCA transport dose-dependently. The PBK model predicted that oral doses of ODE reduced conjugated bile acid levels in plasma. Our simulations match in vivo data and provide a first proof-of-principle for the incorporation of active intestinal bile acid uptake in a bile acid PBK model. This approach could in future be of use to predict the effects of other ASBT-inhibitors on plasma and intestinal bile acid levels.

Bile acids regulate digestion and immune functions. Too little bile acid reuptake in the gut is related to several diseases, including inflammatory bowel disease. This study investigates how reducing bile acid absorption affects bile acid levels in humans using the drug odevixibat (ODE) as an example. ODE reduces bile acid absorption by blocking the intestinal bile acid transporter protein in gut cells. The transport of a bile acid through a gut cell line commonly used to model the intestinal barrier was measured with and without ODE, and mathematical modeling was used to translate the laboratory results to whole-body effects. This combined approach accurately predicted the known effects of ODE on intestinal and bloodstream bile acid levels in humans. This novel approach could be used to predict the effects of other chemicals on intestinal bile acid absorption and intestinal and bloodstream bile acid levels instead of animal testing.

Use of Physiologically Based Kinetic Modeling to Predict Deoxynivalenol Metabolism and Its Role in Intestinal Inflammation and Bile Acid Kinetics in Humans.

Current points of departure used to derive health-based guidance values for deoxynivalenol (DON) are based on studies in laboratory animals. Here, an animal-free testing approach was adopted in which a reverse dosimetry physiologically based kinetic (PBK) modeling is used to predict in vivo dose response curves for DON's effects on intestinal pro-inflammatory cytokine secretion and intestinal bile acid reabsorption in humans from concentration-effect relationships for DON in vitro. The calculated doses for inducing a 5% added effect above the background level (ED5) of DON for increasing IL-1ß secretion in intestinal tissue and for increasing the amounts in the colon lumen of glycochenodeoxycholic acid (GCDCA) were 246 and 36 µg/kg bw/day, respectively. These in vitro-in silico-derived ED5 values were compared to human dietary DON exposure levels, indicating that the risk of DON's effects on these end points occurring in various human populations cannot be excluded. This in vitro-in silico approach provides a novel testing strategy for hazard and risk assessment without using laboratory animals.

Hepatic bile acid synthesis and secretion: Comparison of in vitro methods.

Reliable hepatic in vitro systems are crucial for the safety assessment of xenobiotics. Certain xenobiotics decrease the hepatic bile efflux, which can ultimately result in cholestasis. Preclinical animal models and the currently available in vitro systems poorly predict a xenobiotic's cholestatic potential. Here, we compared the phenotype and capacity of three liver derived in vitro systems to emulate human functionality to synthesize and secrete bile acids (BAs). To this end, basal BA production of sandwich cultured human hepatocytes (SCHHs), HepaRG cells (HepaRGs) and hepatocyte-like intrahepatic cholangiocyte organoids (ICO-heps) were analysed, and the effect of the known BSEP (Bile Salt Export Pump)-inhibitors bosentan and lopinavir on BA disposition in SCHHs and HepaRGs was quantified. RT-qPCR of selected target genes involved in maturation status, synthesis, transport and conjugation of BAs was performed to mechanistically underpin the observed differences in BA homeostasis. ICO-heps produced a (very) low amount of BAs. SCHHs are a powerful tool in cholestasis-testing due to their high basal BA production and high transporter expression compared to the other models tested. HepaRGs were responsive to both selected BSEP-inhibitors and produced a BA profile that is most similar to the human in vivo situation, making them a suitable and practical candidate for cholestasis-testing.

Antibiotic-Induced Changes in Microbiome-Related Metabolites and Bile Acids in Rat Plasma.

Various environmental factors can alter the gut microbiome's composition and functionality, and modulate host health. In this study, the effects of oral and parenteral administration of two poorly bioavailable antibiotics (i.e., vancomycin and streptomycin) on male Wistar Crl/Wi(Han) rats for 28 days were compared to distinguish between microbiome-derived or -associated and systemic changes in the plasma metabolome. The resulting changes in the plasma metabolome were compared to the effects of a third reference compound, roxithromycin, which is readily bioavailable. A community analysis revealed that the oral administration of vancomycin and roxithromycin in particular leads to an altered microbial population. Antibiotic-induced changes depending on the administration routes were observed in plasma metabolite levels. Indole-3-acetic acid (IAA) and hippuric acid (HA) were identified as key metabolites of microbiome modulation, with HA being the most sensitive. Even though large variations in the plasma bile acid pool between and within rats were observed, the change in microbiome community was observed to alter the composition of the bile acid pool, especially by an accumulation of taurine-conjugated primary bile acids. In-depth investigation of the relationship between microbiome variability and their functionality, with emphasis on the bile acid pool, will be necessary to better assess the potential adverseness of environmentally induced microbiome changes.